The objectives are to isolate and characterize antigens associated with human malignant melanoma, to study their specificity, and their possible secretion. If successfully purified, melanoma associated antigens (MAA) will be used to develop an improved assay of humoral immunity to melanoma. The approach will be to iodinate proteins on the surface of melanoma cells by the lactoperoxidase or "Tagit" methods and to label cytoplasmic proteins by incubation with radioactive amino acids. Radiolabelled proteins will be solubilized by lysis with non-ionic detergent and fractionated by column chromatography. MAA will be identified by a double antibody antigen binding assay. The assay is based on the coprecipitation by anti-IgG of complexes of radiolabelled MAA formed with an excess of antimelanoma IgG. Antimelanoma IgG will be prepared from serum of rabbits hyperimmunized to this tumor and exhaustively absorbed with normal human tissue. MAA in antigen containing fractions will be purified and characterized chemically by conventional biochemical procedures. Distribution of MAA in different melanomas will be studied. Specificity of MAA will be studied by quantitative serum absorption, competitive inhibition of antigen binding assay and comparison to radiolabelled proteins in control cells. Control tissues will include normal autologous, homologous, fetal and melanocyte rich tissue, and unrelated malignancies. Possible secretion will be studied by following rate of release of radiolabelled MAA into culture medium. If successfully purified, MAA will be used to develop a sensitive antigen binding assay of the humoral immune response to melanoma. Tumor antigens isolated in this fashion may prove useful in the eventual immunotherapy of this tumor and will facilitate the development of improved assays in the immune response to this cancer. BIBLIOGRAPHIC REFERENCES: Abel, E.A., Bystryn, J-C.: Reproducibility of the immunofluorescent test for anti-melanoma antibodies. J. Invest. Dermat. 66: 117, 1976. Abel, E. A., Bystryn, J-C.: Epidermal cytoplasmic antibodies: incidence and type in normal persons and patients with melanoma. J. Invest. Dermat. 66: 44, 1976.